The highly sensitive in vitro bioassay for measurement of luteinizing hormone (LH) and chorionic gonadotropin (hCG) in blood, developed in our laboratory and termed the RICT assay (rat Leydig cell testosterone bioassay), has been applied to the measurement of serum LH and LH-like gonadotropins in man and several animal species. The high sensitivity of the RICT assays (3-4 fold that of RIA) permits measurements of bioactive LH profiles with frequent sampling in serum from birth to adulthood with good precision. This technique has indicated that the biological activity of LH secreted in man, rhesus monkey and rat is modulated by gonadal steroids (estrogen and androgen). Studies on the pulsatile release of LH in men and postmenopausal women have demonstrated that LH is secreted in pulses of high biological activity as shown by bioimmunoratios significantly increased over interpulse interval. Long-term studies on the in vitro bioactivity of LH and CG in man and other species were continued. The in vitro bioassay has been more extensively applied to studies on gonadotropin regulation in man and other species. We have demonstrated that: 1) when the endogenous GnRH signal is amplified by opiate receptor blockade, bioactive LH is released in more frequent pulses of high biological activity, with a significant increase in androgen production; 2) the RICT is more useful than RIA to follow LH suppression during treatment with GnRH analogs in patients with prostatic carcinoma; 3) a potent gonadotropin-like activity is present in the serum and placenta of pregnant rats, and its secretion is enhanced in the absence of the pituitary gland. Our specific aims are: 1) To define changes in the biochemical properties of pituitary and circulating LH by gonadal steroids and by altered secretion. The ultimate goal is to determine the functional groups of the stored and circulating LH molecule that are responsible for its biological activity. 2) To proceed with further characterization of the newly identified rat chorionic gonadotropin.